Oil and water immersion microscopes have been utilized for many years, and a wide variety of microscope designs using immersion liquids are commercially available. The fundamental features of an immersion microscope are shown in FIG. 1. A specimen 12 to be viewed is mounted on a transparent substrate 14 beneath a transparent coverslip 16. A substage light source/condenser 20 illuminates the specimen 12 through the substrate 14 and the coverslip 16.
An objective lens 24 is positioned above the coverslip 16 and the specimen 12. Light collected by the objective lens 24 passes through additional lenses and optical elements 26 of the microscope, and is sent to an image acquisition system 28 for viewing by a scientist or researcher for example. The image acquisition system 28 may be an eyepiece, CCD camera, or any other system or apparatus used to collect and/or display images of the specimen 12.
Between the objective 24 and the coverslip 16 is an index-matching liquid 30, typically some type of oil, although many different liquids may be used. The liquid 30 advantageously has an index of refraction that matches the coverslip 16 and objective 24, so that no refraction occurs at the interfaces between these components and the liquid 30.
The index matching liquid 30 between the coverslip 16 and the objective 24 increases the amount of light collected by the objective, reduces internal reflections, and improves the overall resolution and image quality produced by the microscope, especially under high magnifying powers.
Prior to the present invention, immersion microscopy had not been performed on specimens at low temperature. Although the need for obtaining magnified images of frozen specimens exists in many applications, the advantages of liquid immersion have never been achieved, due either to freezing of the index matching liquid d or melting of the frozen specimen.